Most hematopoietic cells have a lifespan shorter than that of their host, and senescent cells must continually be replaced in order to maintain homeostasis. Under normal circumstances, the thymus is the primary source for T lymphocyte production. However, the thymus does not contain stem cells for T lymphocytes. Rather, low numbers of bone marrow precursors constantly enter the thymus (via the blood) through venules located deep in the tissue, at the junction of the cortical and medullary compartments (corticomedullary junction, CMJ). These early cells (CD4-8- cells, ~2-3 percent of all thymocytes) then migrate specifically towards the capsule, traversing the cortex (50-100 cell diameters) to accumulate in the sub-capsular zone. After about two days, the polarity of cell migration is reversed. This movement back into the cortex correlates with transition to the CD4+8+ stage of development, and with the bulk of cellular proliferation. Final development requires movement across the CMJ in the opposite direction of early precursors, into the medulla. Thus, thymocytes exhibit a precise pattern of migration during differentiation. In other well-studied models of development, migration into distinct microanatomical sites is known to induce, influence, and/or sustain cellular differentiation and proliferation. The hypothesis put forth in this application is that the differentiation of thymocytes is controlled by enforcing the migration of precursors through the thymus, such that the signals that induce differentiation and proliferation are only delivered in specific microanatomical sites, and for limited periods of time. The aims of this application are to define the zones of the thymus where CD4-8- precursor cells undergo specific differentiative and proliferative changes, and to evaluate the roles of two surface receptors, namely c-kit and the IL-7R, in controlling migration into and out of these zones. A variety of in vivo, ex vivo, and in vitro approaches will be used. These experiments are expected to provide valuable insights into how early T-cell lymphopoiesis and lineage commitment are regulated, and how thymic homeostasis is controlled.